The inferior colliculus (1C)is a central structure in the brainstem auditory pathways and processes almost all ascending auditory sensory information. Acetylcholine is a well-studied neuromodulator and is reported to modulate the responses of cells in the 1C to acoustic stimuli. The vast majority of 1C cells respond to cholinergic agonists, and cholinergic receptors have been localized throughout the 1C. Cholinergic effects contribute to a variety of response properties, including setting a general level of sensitivity as well as altering the selectivity of a cell for properties such as stimulus frequency. There are also suggestions that cholinergic inputs can activate inhibitory circuits in the 1C, but the targets of the cholinergic axons are unknown. Overall, interpreting the results of physiological studies has been hindered by a lack of data on the cholinergic circuitry. Even the source of cholinergic input to the 1C is unknown. Characterizing this circuitry will be an important step in understanding the roles of acetylcholine in auditory function. Multilabel retrograde tracing, immunohistochemistry, and electron microscopy will be used to identify the source(s) of cholinergic input to the 1C and to characterize their collateral projections and synaptic targets Relevance to Public Health: The proposed experiments will fill a large gap in our understanding of cholinergic circuits as they relate to the 1C. A greater understanding of this circuitry will provide a framework for interpreting experiments relating to many aspects of 1C function, as well as experiments related to arousal, sleep/wake cycles, and age- related hearing loss. A better understanding of the circuitry will provide an opportunity to expand our knowledge of the function and dysfunction of the auditory system and will provide insights into the design of new experiments relating to auditory system disorders.